Blood Specimen Collection and Processing

The first step in acquiring a quality lab test result for any patient is the specimen collection procedure. The venipuncture procedure is complex, requiring both knowledge and skill to perform. Several essential steps are required for every successful collection procedure:

Venipuncture Procedure:

• A phlebotomist must have a professional, courteous, and understanding manner in all contact with all patients.
• The first step to the collection is to positively identify the patient by two forms of identification; ask the patient to state and spell his/her name and give you his/her birth date. Check these against the requisition (paper or electronic).
• Check the requisition form for requested tests, other patient information and any special draw requirements. Gather the tubes and supplies that you will need for the draw.
• Position the patient in a chair, or sitting or lying on a bed.
• Wash your hands.
• Select a suitable site for venipuncture, by placing the tourniquet 3 to 4 inches above the selected puncture site on the patient. See below for venipuncture site selection “notes.”
• Do not put the tourniquet on too tightly or leave it on the patient longer than 1 minute.
• Next, put on non-latex gloves, and palpate for a vein.
• When a vein is selected, cleanse the area in a circular motion, beginning at the site and working outward. Allow the area to air dry. After the area is cleansed, it should not be touched or palpated again. If you find it necessary to reevaluate the site by palpation, the area needs to be re-cleansed before the venipuncture is performed.
• Ask the patient to make a fist; avoid “pumping the fist”. Grasp the patient’s arm firmly using your thumb to draw the skin taut and anchor the vein. Swiftly insert the needle through the skin into the lumen of the vein. The needle should form a 15-30 degree angle with the arm surface. Avoid excess probing.

• When the last tube is filling, remove the tourniquet.
• Remove the needle from the patient's arm using a swift backward motion.
• Place gauze immediately on the puncture site. Apply and hold adequate pressure to avoid formation of a hematoma. After holding pressure for 1-2 minutes, tape a fresh piece of gauze or Band-Aid to the puncture site.
• Dispose of contaminated materials/supplies in designated containers.

Note: The larger median cubital and cephalic veins are the usual choice, but the basilic vein on the dorsum of the arm or dorsal hand veins are also acceptable. Foot veins are a last resort because of the higher probability of complications. 

Fingerstick Procedure:

• Follow steps #1 through #5 of the procedure for venipuncture as outlined above.
• The best locations for fingersticks are the 3rd (middle) and 4th (ring) fingers of the non-dominant hand. Do not use the tip of the finger or the center of the finger. Avoid the side of the finger where there is less soft tissue, where vessels and nerves are located, and where the bone is closer to the surface. The 2nd (index) finger tends to have thicker, callused skin. The fifth finger tends to have less soft tissue overlying the bone. Avoid puncturing a finger that is cold or cyanotic, swollen, scarred, or covered with a rash.
• When a site is selected, put on gloves, and cleanse the selected puncture area.
• Massage the finger toward the selected site prior to the puncture.
• Using a sterile safety lancet, make a skin puncture just off the center of the finger pad. The puncture should be made perpendicular to the ridges of the fingerprint so that the drop of blood does not run down the ridges.
• Wipe away the first drop of blood, which tends to contain excess tissue fluid.

• Collect drops of blood into the collection tube/device by gentle pressure on the finger. Avoid excessive pressure or “milking” that may squeeze tissue fluid into the drop of blood.
•  Cap, rotate and invert the collection device to mix the blood collected.
• Have the patient hold a small gauze pad over the puncture site for a few minutes to stop the bleeding.
• Dispose of contaminated materials/supplies in designated containers.
• Label all appropriate tubes at the patient bedside.

Heelstick Procedure (infants):

• The recommended location for blood collection on a newborn baby or infant is the heel. The diagram below indicates the proper area to use for heel punctures for blood collection.

• Prewarming the infant's heel (42° C for 3 to 5 minutes) is important to increase the flow of blood for collection.
• Wash your hands, and put gloves on. Clean the site to be punctured with an alcohol sponge. Dry the cleaned area with a dry gauze pad.
• Hold the baby's foot firmly to avoid sudden movement.
• Using a sterile blood safety lancet, puncture the side of the heel in the appropriate regions shown above. Make the cut across the heel print lines so that a drop of blood can well up and not run down along the lines.
• Wipe away the first drop of blood with a piece of clean, dry cotton gauze. Since newborns do not often bleed immediately, use gentle pressure to produce a rounded drop of blood. Do not use excessive pressure because the blood may become diluted with tissue fluid.
• Fill the required microtainer(s) as needed.
• When finished, elevate the heel, place a piece of clean, dry cotton on the puncture site, and hold it in place until the bleeding has stopped.  Apply tape or Band-Aid to area if needed.
• Be sure to dispose of the lancet in the appropriate sharps container. Dispose of contaminated materials in appropriate waste receptacles.
• Remove your gloves and wash your hands.

Order of Draw:

Blood collection tubes must be drawn in a specific order to avoid cross-contamination of additives between tubes. The recommended order of draw for plastic vacutainer tubes is:

    1.  First - blood culture bottle or tube (yellow or yellow-black top)
    2. Second - coagulation tube (light blue top).
    3. Third - non-additive tube (red top)
    4. Last draw - additive tubes in this order:

•  SST (red-gray or gold top). Contains a gel separator and clot activator.
•  Sodium heparin (dark green top)
•  PST (light green top). Contains lithium heparin anticoagulant and a gel separator.
•  EDTA (lavender top)
•  Oxalate/fluoride (light gray top) or other additives

NOTE: Tubes with additives must be thoroughly mixed. Clotting or erroneous test results may be obtained when the blood is not thoroughly mixed with the additive.

Labeling The Sample

A properly labeled sample is essential so that the results of the test match the patient. The key elements in labeling are:

• Patient's surname, first and middle names.
• Patient's ID number. NOTE: Both of the above MUST match the information on the requisition form.
• Date, time and initials of the phlebotomist must be on the label of EACH tube or electronically entered. Automated systems may include labels with bar codes.

Areas to Avoid When Choosing a Site for Blood Draw:

Certain areas are to be avoided when choosing a site for blood draw:

• Extensive scars from burns and surgery - it is difficult to puncture the scar tissue and obtain a specimen.
• The upper extremity on the side of a previous mastectomy - test results may be affected because of lymphedema.
• Hematoma - may cause erroneous test results. If another site is not available, collect the specimen distal to the hematoma.
• Intravenous therapy (IV) / blood transfusions - fluid may dilute the specimen, so collect from the opposite arm if possible.
• Cannula/fistula/heparin lock - hospitals have special policies regarding these devices. In general, blood should not be drawn from an arm with a fistula or cannula without consulting the attending physician.
• Edematous extremities - tissue fluid accumulation alters test results.

Techniques to Prevent Hemolysis (which can interfere with many tests):

• Mix all tubes with anticoagulant additives gently (vigorous shaking can cause hemolysis) 5-10 times.
• Avoid drawing blood from a hematoma; select another draw site.
• If using a needle and syringe, avoid drawing the plunger back too forcefully.
• Make sure the venipuncture site is dry before proceeding with draw.
• Avoid a probing, traumatic venipuncture.
• Avoid prolonged tourniquet application (no more than 2 minutes; less than 1 minute is optimal).
• Avoid massaging, squeezing, or probing a site.
• Avoid excessive fist clenching.
• If blood flow into tube slows, adjust needle position to remain in the center of the lumen.

Blood Sample Handling and Processing:

Pre-centrifugation Handling - The first critical step in the lab testing process, after obtaining the sample, is the preparation of the blood samples. Specimen integrity can be maintained by following some basic handling processes:

• Fill tubes to the stated draw volume to ensure the proper blood-to-additive ratio. Allow the tubes to fill until the vacuum is exhausted and blood flow ceases.
• Vacutainer tubes should be stored at 4-25°C (39-77°F).
• Tubes should not be used beyond the designated expiration date.
• Mix all gel barrier and additive tubes by gentle inversion 5 to10 times immediately after the draw. This assists in the clotting process. This also assures homogenous mixing of the additives with the blood in all types of additive tubes.
• Serum separator tubes should clot for a full 30 minutes in a vertical position prior to centrifugation. Short clotting times can result in fibrin formation, which may interfere with complete gel barrier formation.

Blood Sample Centrifugation – It is recommended that serum be physically separated from contact with cells as soon as possible, with a maximum time limit of 2 hours from the time of collection.

• Complete gel barrier formation (gel barrier tubes) is time, temperature and G-force dependent. The uniformity of the barrier is time dependent; an incomplete barrier could result from shortened centrifugation times.

• In general, for a horizontal, swing-bucket centrifuge, the recommended spin time is 10 minutes. For a fixed-angle centrifuge, the recommended spin time is 15 minutes.
• NOTE: Gel flow may be impeded if chilled before or after centrifugation.
• Tubes tops should remain closed at all times during the centrifugation process.

• Place the closed tubes in the centrifuge as a “balanced load” noting the following:

1. Opposing tube holders must be identical and contain the same cushion or none at all.
2. Opposing tube holders must be empty or loaded with equally weighted samples (tubes same size and equal in fill).
3. If an odd number of samples is to be spun, fill a tube with water to match the weight of the unpaired sample and place it across from this sample.

Centrifuge Safety

• Interference with an activated centrifuge by an impatient employee can result in bodily injury in the form of direct trauma or aerosolization of hazardous droplets.
• Centrifuges must never be operated without a cover in place.
• Uncovered specimen tubes must not be centrifuged.
• Centrifuges must never be slowed down or stopped by grasping part(s) of the device with your hand or by applying another object against the rotating equipment.
• Be sure the centrifuge is appropriately balanced before activating. If an abnormal noise, vibration, or sound is noted while the centrifuge is in operation, immediately stop the unit (turn off the switch) and check for a possible load imbalance.
• Clean the centrifuge daily with a disinfectant and paper towel. Broken tubes or liquid spills must be cleaned immediately.

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